Sulphur isotope analysis (?34S) is increasingly identified as a valuable tool for source differentiation and the determination of trophic level in food webs, but there are still many uncertainties associated with the interpretation of d34S data. To investigate the effects of temperature, ration, body size and age on sulphur trophic fractionation (?d34S) in fish, we reared European sea bass (Dicentrarchus labrax) on identical diets at 11 and 16°C at three ration levels for over 600 days. ?d34S was between 0 and -1‰. The effect of temperature on ?d34S was small and inconsistent, varying over the course of the experiment and depending on ration. This contrasts with temperature effects on bass ?d 13C and ?d15N, where ?d 13C increases at warm temperatures while ?d15N falls. Body size and age had a positive relationship with ?d 34S but the relationship with size was not significant for bass that weighed >20 g. As ?d34S is small and the range in d34S of potential diet items can be much greater than the range in d13C or d15N, our results show that sulphur stable isotopes are particularly useful for source differentiation in fish.